Heparin and heparan sulfate are glycosaminoglycans having a repeating unit (4GlcAβ1/IdoAα1→4GlcNAcα1→) composed of two sugars, i.e., a hexuronic acid (HexA) residue (D-glucuronic acid (GlcA) or L-iduronic acid (IdoA) residue) and an N-acetylglucosamine (GlcNAc) residue as a basic skeleton (heparin skeleton), basically with a portion of hydroxyl at position 2 of the hexuronic acid residues and a portion of amino groups at position 2 or hydroxyl at position 6 of the N-acetylglucosamine residue being replaced with sulfates.
Cloning of the gene of an enzyme which transfers sulfate to glycosaminoglycans (glycosaminoglycan sulfotransferase) has made it possible to readily obtain the enzyme in amounts sufficient enough to obtain information on the biosynthesis of glycosaminoglycans having sulfates (sulfated glycosaminoglycans). This will presumably provide a useful approach to investigation on the relationship between the structure and function of sulfated glycosaminoglycans. It has been known that the biosynthesis of sulfated glycosaminoglycans, in particular, biosynthesis of heparin and heparan sulfate, is achieved by a variety of sulfation processes (Kobata, H., Hakomori, S., Nagai, K., Glycotechnology (5), 57 (1994), published by Kodansha Scientific). This sulfation may involve various glycosaminoglycan sulfotransferases. As the glycosaminoglycan sulfotransferases which transfer-sulfate to heparin or heparan sulfate, heparan sulfate N-sulfotransferase (hereinafter in some cases abbreviated as “HSNST”) which catalyzes de-N-acetylation of N-acetylglucosamine residue and sulfate transfer; heparan sulfate 2-O-sulfotransferase (hereinafter in some cases abbreviated as “HS2ST”) which transfers sulfate to hydroxyl at position 2 of hexuronic acid residue; and heparan sulfate 6-O-sulfotransferase (hereinafter in some cases abbreviated as “HS6ST”) which transfers sulfate to hydroxyl at position 6 of N-sulfated glycosamine residue, have been isolated and for some of the sulfotransferases, cDNA cloning has already been performed.
The inventors of the present invention have already purified HS6ST which selectively transfers sulfate from 3′-phosphoadenosine 5′-phosphosulfate which is the sulphate donor, to hydroxyl at position 6 of N-sulfated glucosamine residue contained in heparan sulfate which is the sulfate acceptor, from cultured cells derived from Chinese hamster, mouse, and human (J. Biol. Chem., 270, 4172-4179 (1995)), have completed cloning of the enzyme (J. Biol. Chem., 273, 9208-9213 (1998)), and have succeeded in further cloning two of the isoforms in mouse (J. Biol. Chem., 275, 2859-2868 (2000)).